We propose to develop rapid-scanning optical-resolution photoacoustic microscopy (OR-PAM), which is capable of label- free imaging of non-fluorescent optical absorbers. Optical absorption contrast is highly desirable for microvascular imaging and characterization because of the presence of endogenous strongly light-absorbing hemoglobin. The proposed imaging system can provide a 5-micron lateral resolution, a 15-micron axial resolution, a 1.2-mm imaging depth, and a 30-Hz B-scan frame rate. The following specific aims are proposed. Aim 1. System design and implementation: The proposed rapid-scanning OR-PAM is expected to outperform the current version in imaging speed by a factor of 10. Nearly diffraction-limited optical focusing is employed to achieve the lateral resolution. Absorption of the focused light produced photoacoustic waves due to thermoelastic expansion. Detection of the photoacoustic waves using a high-frequency focused ultrasonic transducer measures the internal light absorption distribution and hence provides a 3D mapping of the optical structure. OR-PAM is able to measure optical absorption with a relative sensitivity of 100%, the theoretical limit of any linear-effect imaging methods. Rapid scanning of the dual optical-ultrasonic foci is required to achieve a high imaging rate. Aim 2. Testing the imaging system in phantoms and living mice: The proposed OR-PAM system will be tested with phantoms and living mice to quantify the maximum imaging depth, spatial resolution, SNR, and the frame rate for B- scan imaging. PUBLIC HEALTH RELEVANCE: Microcirculation, the distal functional unit of the cardiovascular system, provides exchange sites for gases, nutrients, metabolic wastes, and thermal energy between the blood and the tissues. Pathologic microcirculation reflects the breakdown of homeostasis in organisms, which ultimately leads to tissue inviability. Therefore, the proposed in vivo microvascular imaging and characterization is of significant physiological, pathophysiological, and clinical importance.